1. Field of the Invention
This invention relates to diagnostic ultrasonic imaging and contrast agents for use thereof. More particularly, it relates to ultrasonic contrast agents comprising emulsions capable of forming gas microbubbles upon the application of ultrasonic energy and methods for their use in diagnostic imaging.
2. Brief Description of the Background Art
Diagnostic ultrasonic imaging is based on the principal that waves of sound energy can be focused upon an area of interest and reflected in such a way as to produce an image thereof. The ultrasonic scanner utilized is placed on a body surface overlying the area to be imaged, and sound waves are directed toward that area. The scanner detects reflected sound waves and translates the data into images. When ultrasonic energy is transmitted through a substance, the amount of energy reflected depends upon the velocity of the transmission and the acoustic properties of the substance. Changes in the substance's acoustic properties (e.g. variations in acoustic impedance) are most prominent at the interfaces of different substances, such as liquid-solid or liquid-gas. Consequently, when ultrasonic energy is directed through media, changes in acoustic properties will result in more intense sound reflection signals for detection by the ultrasonic scanner.
Ultrasonic imaging agents of particular importance are composed of gas-containing substances which, when injected into the circulatory system, provide improved sound reflection and image clarity. One class of gas-containing imaging agents consists of microspheres of gas surrounded by a shell made of a biocompatible substance. These are best typified by ALBUNEX.RTM. (Molecular Biosystems, San Diego, Calif.: U.S. Pat. Nos. 4,572,203; 4,718,433; 4,744,958; 4,844,882 and 4,957,656) which consists of microspheres of air surrounded by albumin shells. Another such microspheric imaging agent is described by Holmes, et al. These microspheres consist of either non-proteinaceous crosslinked or polymerized amphipathic moieties forming micelles (PCT WO 92/17212) or crosslinked proteins (PCT WO 92/17213), both of which encapsulate gasses such as nitrogen, SF.sub.6 and CF.sub.4.
Another class of ultrasonic imaging agents can be described as microparticles of a solid or semi-solid substance containing gas which is entrapped in the microparticle matrix during production. Glajich, et al. (U.S. Pat. No. 5,147,631) describe the formation of porous particles of an inorganic material containing entrapped gas or liquid such as O.sub.2, CF.sub.4, perfluoroethane and argon. Erbel, et al. (U.S. Pat. No. 5,137,928) describe polyamino-dicarboxylic acid-co-imide derivatives capable of entrapping gasses such as air, argon and krypton. Albayrak, et al. (European Patent Specification 0 357 163) describe crystalline complexes entrapping gasses such as nitrogen, krypton, SF.sub.6, cyclopropane and pentane which are dissolved in an aqueous vehicle such as protein or glycerol causing the release of gas bubbles. The aqueous vehicle, now containing a plurality of microbubbles of gas in solution, is then ready for use as an injectable ultrasonic imaging agent. Stein, et al. (European Patent Specification 327 490) describe microparticles containing amyloses or synthetic biodegradable polymers entrapping gasses or liquids with a boiling point less than 60.degree. C.
Another class of gas-containing imaging agents are lipid vesicles or liposomes. Unger (U.S. Pat. Nos. 5,088,499 and 5,123,414) describes the encapsulation of gasses or gaseous precursors in liposomes, more particularly liposomes which contain ionophores for activation of gaseous precursors by way of a pH gradient. Henderson, et al. (PCT WO 92/15824) describe lipid vesicles with gas-filled center cores.
Still another class of imaging agents is composed of microbubbles of gas in solution. For example, Tickner, et al. (U.S. Pat. No. 4,276,885) describe microbubbles dispersed in liquified gelatin. More-recently, Quay (PCT WO 93/05819) describes ultrasound imaging agents comprising microbubbles of selected gasses in solution. In a specific embodiment, Quay describes the formation of a gas-liquid emulsion of decafluorobutane. Also disclosed therein are imaging agents comprising aqueous dispersion of biocompatible gasses, some of which are gaseous at ambient temperature and others of which become gaseous at the body temperature of the subject being imaged.
The efficiency of gas as an ultrasound imaging agent is described by J. Ophir and K. J. Parker, Contrast Agents in Diagnostic Ultrasound, Ultrasound in Medicine and Biology (1989), Vol. 15(4) p. 319-333. However, the disadvantages of using gas as an ultrasound imaging agent have been and continue to be lacking of sufficient persistence of the gas in-vivo and in-vitro, and toxicity due to the introduction of gas into the venous system.
The present invention relates to site specific oil-in-water emulsions and is based on the unexpected observation that emulsions of gas-forming chemicals can be stabilized in the liquid state and will produce microbubbles when subjected to ultrasonic energy. The advantages are that such emulsions are more stable than most of the gas-containing imaging agents heretofor described, and their ability to form microbubbles when subjected to ultrasonic energy makes them site-specific and inherently less toxic due to less overall gas being introduced into the venous system.